The roles of mDia2 in hematopoietic stem and progenitor cell engraftment and migration

项目摘要 植入失败是造血干细胞移植的主要局限性之一， 用于治疗许多良性和恶性血液病。除了免疫介导的排斥反应外， 移植的细胞，造血干细胞的无效迁移、粘附和植入， 造血祖细胞（HSPCs）向骨髓小生境的迁移是移植失败的重要原因。的 调节HSPC植入和迁移至骨髓的细胞内信号传导途径通常直接 或通过调节肌动蛋白聚合的蛋白质间接靶向肌动蛋白细胞骨架网络。mDia formin是一个主要的肌动蛋白成核家族，但其在HSPCs中的功能尚不清楚。我们的初步研究 使用我们最近产生的mDia 2造血特异性敲除小鼠的研究表明，mDia 2的缺失导致 在竞争性移植试验中，HSPC粘附和骨髓植入存在显著缺陷。 mDia 2的缺失也显著降低了G-CSF诱导的HSPC从骨髓中的动员。我们也 发现β 2整合素包括CD 11 a/CD 18（LFA 1）和CD 11b/CD 18（Mac 1）在CD 11 a/CD 18（LFA 1）和CD 11b/CD 18（Mac 1）中显著表达。 随着mDia 2的丢失而下调。这些结果使我们假设mDia 2对于 造血干细胞和祖细胞的植入、粘附和迁移。我们提出了三个 具体目的是检验这一假设。首先，我们将确定mDia 2在HSPC中的功能作用 植入、粘附和迁移至骨髓。具体来说，我们将研究mDia 2在 HSPCs在体内外的迁移、粘附及其与骨髓生态位的关系。 我们还将在体内使用mDia激活剂，以探索mDia激活的临床前模型，用于增强干细胞 细胞动员和植入。在这些实验中，相对纯的SLAM+造血干细胞 人口也将受到考验。其次，我们将研究mDia 2在调控HSPC中的作用机制， 通过新的mDia 2-MAL-SRF-LFA 1/Mac 1途径的移植、粘附和迁移。三是立足 我们的初步研究表明，乙酰化mDia 2影响其在肌动蛋白聚合的活性，我们将确定 mDia 2的乙酰化或其他翻译后修饰是否以及如何影响其在HSPC中的功能 移入和迁移。这项拟议研究的总体目标是了解mDia 2在以下方面的作用： HSPC植入、粘附和迁移并使用体内小鼠模型阐明其机制， 移植试验更好地了解这些过程可能会导致识别新的 造血干细胞移植治疗策略的靶点。

PROJECT SUMMARY Failure in engraftment is one of the major limitations of hematopoietic stem cell transplantation, which is critical for the treatment of many benign and malignant hematologic disorders. Besides immune-mediated rejection of the transplanted cells, ineffective migration, adhesion, and engraftment of the hematopoietic stem and progenitor cells (HSPCs) to the bone marrow niche are important causes of engraftment failure. The intracellular signaling pathways regulating HSPC engraftment and migration to the bone marrow often directly or indirectly target the actin cytoskeleton network through proteins that regulate actin polymerization. The mDia formins are a major actin-nucleating family, but their functions in HSPCs are unknown. Our preliminary studies using our recently generated mDia2 hematopoietic-specific knockout mice showed that loss of mDia2 resulted in significant defects in HSPC adhesion and bone marrow engraftment in competitive transplantation assays. Loss of mDia2 also markedly reduced G-CSF-induced HSPC mobilization out of the bone marrow. We also found that beta2 integrins including CD11a/CD18 (LFA1) and CD11b/CD18 (Mac1) were significantly downregulated with the loss of mDia2. These results led us to hypothesize that mDia2 is critical for the engraftment, adhesion, and migration of the hematopoietic stem and progenitor cells. We proposed three specific aims to test this hypothesis. First, we will determine the functional roles of mDia2 in HSPC engraftment, adhesion, and migration to the bone marrow. Specifically, we will investigate the role of mDia2 in the in vitro and in vivo migration and adhesion of HSPCs and their relationship with the bone marrow niche. We will also use an mDia activator in vivo to explore a pre-clinical model of mDia activation for enhancing stem cell mobilization and engraftment. In these experiments, relatively pure SLAM+ hematopoietic stem cell population will also be tested. Second, we will determine the mechanisms of mDia2 in regulating HSPC engraftment, adhesion, and migration through a novel mDia2-MAL-SRF-LFA1/Mac1 pathway. Third, based on our preliminary studies that acetylation of mDia2 affects its activity in actin polymerization, we will determine whether and how acetylation or other post-translational modifications of mDia2 influence its functions in HSPC engraftment and migration. The overall goal of this proposed study is to understand the roles of mDia2 in HSPC engraftment, adhesion, and migration and elucidate its mechanisms using in vivo mouse models and transplantation assays. A better understanding of these processes may lead to the identification of novel targets for therapeutic strategies in hematopoietic stem cell transplantation.